Spermatid Translocation in the Rat Seminiferous Epithelium: Coupling Membrane Trafficking Machinery to a Junction Plaque1

Abstract

In this study, we demonstrate that specialized junction plaques that occur between Sertoli cells and spermatids in the rat testis support microtubule translocation in vitro. During spermatogenesis, Sertoli cells are attached to spermatids by specialized adhesion junctions termed ectoplasmic specializations (ESs). These structures consist of regions of the plasma membrane adherent to the spermatid head, a submembrane layer of tightly packed actin filaments, and an attached cistern of endoplasmic reticulum. It has been proposed that motor proteins on the endoplasmic reticulum interact with adjacent microtubules to translocate the junction plaques, and hence the attached spermatids, within the epithelium. If this hypothesis is true, then isolated junctions should support microtubule transport. To verify this prediction, we have mechanically isolated rat spermatids, together with their attached ESs, and tested them for their ability to transport microtubules in vitro. Most assays were done in the presence of 2 mg/ml testicular cytosol and at room temperature. ESs attached to spermatids supported microtubule translocation. In some cases in which motility events were detected, microtubules moved smoothly over the junction site. In others, the movement was slow but progressive, saltatory and "inch-worm-like." No motility was detected in the absence of exogenous ATP or in the presence of apyrase (an enzyme that catalyses the breakdown of ATP). Our results are consistent with the microtubule-based motility hypothesis of spermatid translocation.

A Kinesin Is Present at Unique Sertoli/Spermatid Adherens Junctions in Rat and Mouse Testes1

“…Microtubules bind to ectoplasmic specializations attached to spermatids mechanically separated from the epithelium, and the binding is ATP dependent [31]. In motility assays, also using isolated spermatids with attached junctions, ectoplasmic specializations are capable of transporting microtubules [36]. Significantly, microtubules move both in the plus end direction and in the minus end direction across ectoplasmic specializations [9], a result consistent with the presence of at least two types of motor proteins on the structures and with the down and up directions of spermatid translocation in vivo.…”

Cytoskeletal dynamics and spermatogenesis

“…Early studies demonstrated that the intratesticular injection of taxol (an MT stabilizer) led to the retention of step 19 spermatids deep within Sertoli cells crypts, instead of being transported towards the tubule lumen (Russell et al 1989). Indeed, when mechanically isolated spermatids which retained their associated ES from the Sertoli cell were immobilized in a flow chamber, movement of fluorescently labelled MTs across the spermatid surface was observed in the presence of exogenous ATP (Beach & Vogl 1999). This illustrated that a force can be generated between ES-bearing spermatids and MTs that causes their movement in opposite directions.…”

Parallel actin bundles and their multiple actin-bundling proteins

“…The parallel actin bundles of the ectoplasmic specialization junctional plaque are believed to function in part as a scaffold that supports and stabilizes an adhesive domain in the Sertoli cell plasma membrane [44,45]. They are also believed to act indirectly, via their connection to the cistern of endoplasmic reticulum, as a link to an underlying network of microtubules that may be responsible for changes in the depth of the ectoplasmic specialization-spermatid complex within the seminiferous epithelium [45,46]. Espin, the founding member of the espin family of actin-bundling proteins, was identified as a component of the parallel actin bundles in the ectoplasmic specialization junctional plaque …”